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Greek colonisation of South Italy and Sicily (Magna Graecia) was a defining event in European cultural history, although the demographic processes and genetic impacts involved have not been systematically investigated. Here, we combine high-resolution surveys of the variability at the uni-parentally inherited Y chromosome and mitochondrial DNA in selected samples of putative source and recipient populations with forward-in-time simulations of alternative demographic models to detect signatures of that impact. Using a subset of haplotypes chosen to represent historical sources, we recover a clear signature of Greek ancestry in East Sicily compatible with the settlement from Euboea during the Archaic Period (eighth to fifth century BCE). We inferred moderate sex-bias in the numbers of individuals involved in the colonisation: a few thousand breeding men and a few hundred breeding women were the estimated number of migrants. Last, we demonstrate that studies aimed at quantifying Hellenic genetic flow by the proportion of specific lineages surviving in present-day populations may be misleading.A total of 59 previously published single nucleotide polymorphisms was analysed following a hierarchical genotyping strategy. Samples were amplified in a standard PCR reaction and the SNaPshot Multiplex System (Life Technologies Corp., Carlsbad, CA, USA) primer extension protocol was used. All samples were first genotyped for markers, E-M35, F-M89, G-M201, H-M282, I-M170, K-M9, J-M172, J-M267, J-M304, R-M173, P-M45, R-M17 and R-M269, to classify them into major European branches. Samples belonging to haplogroups E-M35, E-M78, J-M172, I-M170 and R-M269 were further analysed by means of haplogroup specific multiplexes (Supplementary Table S2). Furthermore, samples assigned to haplogroup G-M201 were analysed for markers M406 and P15 through direct sequencing (Supplementary Table S3). Nomenclature used for haplogroup labelling follows YCC conventions26 and recent updating (ISOGG Y-Tree 2015 http://www.isogg.org/tree/).
The entire data set was also analysed at a total of 26 Y chromosome short tandem repeats (YSTRs): those included in the AmpFlSTR Yfiler PCR Amplification kit (Applied Biosystems, Foster City, CA) and additional 9 YSTRs (DYS460, DYS388, YCA-II a/b, DYS461, DYS445, YGATA-A.10 and DYS413 a/b) by suitably designed multiplexed-PCR reactions (Supplementary Table S4). We finally assembled a haplotype data set based on 20 of the 26 analysed STR markers by excluding those STRs that in the PCR analysis co-amplify two loci and whose allele assignation to a defined locus was not possible (DYS385 a/b, YCA-II a/b and DYS413 a/b). A subset of samples (N=304) has been analysed for the hypervariable region I of mitochondrial DNA (mtDNA) using primers 15997L and 017H.Population relationships
Y haplogroup frequencies are reported in Supplementary Table S7 while the overall pattern of inter-population genetic relationships is shown in Figure 2 and Supplementary Table S10. Cretan, mainland Greek and Lebanese samples were introduced to widen the spectrum of the historical players acting in south-eastern Mediterranean at the time of the GC, as proxies of non-Corinthian Dorian colonisers of South Sicily (since Crete contributed to the foundation of Gela and, in turn, Akragas), non-Euboean Ionian colonisers of South Italy and East Sicily, and Phoenician settlers in West Sicily, respectively. Looking at the reciprocal positions on the plot, little evidence of these historical events emerges, with the positioning reflecting geography rather than history. Accordingly, we observe higher genetic distance than that expected based on archaeological evidence between putative descendants of source (Greeks from Ionia, Corinthians and Cretan) and recipient (Sicilians) groups of the GC, as well as between the putative founders (Lebanese) of the Phoenician colonies in western Sicily (Motya, Panormos and Solus) and the present-day population.Signatures of the Archaic Hellenic contribution
To detect genetic signatures of Greek migration in southern Italy and Sicily related compatible with the Archaic scenario, we compared fractions of haplotype pairs within the 8–12 mutational range, or 'GChp', with the same fractions obtained by using Albanian, Croatian and Turkish samples as reference sourcesSamples from East Sicily, West Sicily, South and Central Italy showed significant (P<0.01) enrichment of GChps when the Greek sample from the Euboea Island was compared with Corinthia and reference sources. Except in West Sicily and Central Italy, this enrichment remained highly significant even after correcting for multiple tests (P<0.05).41 Conversely, when considering Corinthia against other reference sources, none of the recipient samples showed a full set of significant values. The comparisons involving recipients versus Albania most commonly showed a lack of significant enrichment in GChps with respect to Euboea and Corinthia. We reasoned that contacts either between sources and recipients or between sources after the GC, that is, during the Classical and Christian periods, might have contributed to increase the GChps rate. Thus, to provide more stringent conditions for haplotype identification, we excluded all haplotypes with a molecular distance less than seven mutational steps. This 'filtered' data set confirmed the pattern observed with the less stringent criteria for the East Sicily/Euboea pair, which showed significant enrichment in GChps in two out of three comparisons even after the Bonferroni correction (Table 1b). The results for West Sicily and South Italy did not hold statistical significance when a Bonferroni correction was applied. None of the other Italian recipients showed a full set of significant enrichment with Euboea or Corinthia.Estimating Greek contribution
The number of GChps identified using the suggested molecular distance cannot be used either to directly estimate the current Greek legacy in Italian populations or to provide an indication of the original demographic contribution. Nevertheless, this approach helped us to identify populations (East Sicily, and, to a lesser degree, West Sicily and South Italy), that are characterized by a significant association with Greek populations derived during the time window of interest.http://www.nature.com/ejhg/journal/v...g2015124a.htmlTo quantify the original demographic impact of the Greek settlers inferable from present-day Y chromosome variability, we explored two main census scenarios using a simulation-based approach. In the first scenario, high count, we based our model on the demographic estimates of Beloch,18, 19 who suggested a census size of 1.35 million people for Sicily and of 3 million for Greece at the time of the Hellenic colonisation in the Archaic Period. In the second scenario, low count, we modelled population size estimates that were smaller by an order of magnitude.21 Nevertheless, the two scenarios have similar source/recipient effective size ratios (S/R). If we assume that the proportions of past Sicilian, Euboean and Peloponnese census and male (and female) effective population sizes is one-sixth of the current census size, we estimate a S/R of 3.65 for the high-count model and a S/R of 3.75 for the low-count model. Simulation results are reported on Figure 3 and Supplementary Table S8. When considering Y-STR haplotypes, the observed DHS value between Euboea and East Sicily (0.5353) is compatible with an effective number of migrants ranging between 500 and 5000, clearly rejecting larger contributions (10 000), irrespective of the scenario considered. The DHS value obtained for hypervariable region I haplotypes (0.5995) supports an effective number of migrants between 500 and 1000, with larger contributions clearly excluded. When the NRY- and mtDNA-based estimates are paired according to the demographic model, the male-to-female migrant ratio ranged between 1:1 and 2:1 under a population growth model and between 2:1 and 10:1 under a constant population size model.
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